1. Field of the Invention
The present invention relates to an optical device such as a microscope system and an imaging method that can correctly generate a single image without incongruity from a plurality of divided observed images.
Priority is claimed on Japanese Patent Application No. 2003-401429, filed Dec. 1, 2003, the content of which is incorporated herein by reference, and Japanese Patent Application No. 2004-101603, filed Mar. 31, 2004, the content of which is incorporated herein by reference.
2. Description of Related Art
Gene sequences of many living organisms, including humans, are now known due to recent advances in gene sequencing technology. In addition, the causal connections between gene products such as sequenced proteins and diseases are slowly beginning to be understood. Furthermore, various visualization methods and devices employing cells are beginning to be conceived for exhaustive and statistical analysis of proteins, genes, etc. In particular, to perform such analysis, it is necessary to obtain desired information while cultivating cells over a long period. Doing so requires an apparatus that allows incubation and observation of cells under a microscope.
One such apparatus is known that uses a transparent thermostatic incubator for microscopy (for example, see paragraphs 0004 to 0007 and FIGS. 1 to 4 of Japanese Unexamined Patent Application, First Publication No. H10-28576).
This transparent thermostatic incubator for microscopy (hereafter, simply referred to as an incubator) is constituted to allow the setting of various incubation conditions for cells. For this purpose, the incubator has a pair of transparent exothermic plates, a carbon dioxide supply port and a carbon dioxide exhaust port, and an evaporation dish. Here, the temperatures of the transparent exothermic plates can be controlled to predetermined values by a thermostat. The carbon dioxide supply port and exhaust port control the carbon dioxide gas concentration in the space. The evaporation dish maintains the humidity in the hermetically sealed container.
In observations using the incubator, the temperature, carbon dioxide gas concentration and humidity in the container can be controlled, thereby enabling observation during cell culturing. Specifically, changes in the cell incubation state over time can be observed, for example, from the lower side of the transparent exothermic plates by observation with an objective lens.
Another cell observation method is fluorescence imaging. In particular, the liposome method, gene gun method and microinjection method, etc., are methods of observing fluorescence in a living material.
With these methods, technology for observing dynamic changes in cells and the like in which genes have been introduced has made rapid strides. In recent years, with the striking progress of green fluorescent protein, techniques of continuous fluorescence observation under a microscope of temporal and spatial changes in the subject of observation are attracting attention. The aforementioned subject of observation includes various behavior of materials in organelles in living cells and proteins.
When employing such observation methods, the need arises for high-resolution and wide-field observation of fluorescence from a microarray. This microarray comprises a large number of spots, ranging from several hundred to several tens of thousands, on a plastic or glass dish, plate or slide glass. During observation, this microarray is disposed in the incubator.
When observing a specimen with a microscope, the field that can be observed at one time is primarily determined by the magnification of the objective lens. Therefore, the higher the magnification of the objective lens, the narrower the field of view, which restricts the observed field to only a small portion of the sample. Until now, for example, microscopic image outputting methods (for example, see page 3, upper right column, line 5, to page 6, upper left column, line 6, and FIGS. 1 to 4 of Japanese Unexamined Patent Application, First Publication No. H03-209415) and methods based on microscopic systems (for example, see sections 0015 to 0087, and FIGS. 1 to 13 of Japanese Unexamined Patent Application, First Publication No. H09-281405) have been known as methods for obtaining a high resolution and wide field microscopic image.
In these microscopic image outputting methods, scanning is performed by changing the relative positional relationship between the stage on which the specimen is mounted and the illumination, and partial images dividing the imaging region into a plurality of regions are recorded. Then, the partial images are composited so as to be tiled to produce an image of the entire specimen.
Methods based on microscope systems are methods of reconstructing an entire image of a specimen by image compositing. First, the entire specimen is divided into small regions so as to have a plurality of mutually overlapping portions. Then, the image of each small region is captured with a microscope, and detection of positional deviations is performed by comparing the overlapping portions of each captured image. The images are then composited after correcting the position of the image of each small region based on these positional deviations. In this way, a wide-field high-definition image of the specimen is produced.